Bioscaffold engineering has the potential to vastly improve the treatment of tissue injuries, especially those associated with tumor, trauma, and congenital deficiencies where autograft tissue (i.e., tissue harvested from the recipient patient) or allograft tissue (i.e., tissue harvested from a different subject of the same species) might not be available in sufficient quantity for reconstruction. Current tissue-engineering strategies have relied on scaffolds derived from both synthetic (e.g., polyglycolic acid) and naturally-derived (e.g., collagen) materials to form the cell-scaffold construct. Currently available tissue scaffold products include small intestine submucosa (Restore™, porcine SIS, DePuy Orthopaedics), (CuffPatch™, porcine SIS, Organogenesis), (SIS; Cook Biotech, Inc.), reformulated collagen scaffolds (3D Collagen Composite, BD Biosciences), acellular human dermal collagen matrices (Graftjacket®, Wright Medical Technologies), fetal bovine dermis (TissueMend®, Stryker), and synthetic polymer scaffolds, primarily polyesters (e.g. PGA, PCL, and PLA).
Synthetic scaffolds can produce breakdown products that have been shown to be antimitotic and cytotoxic in vivo (Garvin et al. (2003) Tissue Eng 9(5):967-979). Many synthetic and naturally-derived scaffolds also lack the initial mechanical strength to permit immediate motion and rehabilitation after implantation, leading to subsequent adhesion formation, decreased range of motion, and poor functional outcomes (Cao et al. (2002) Plast Reconstr Surg 110:1280-1289; Buckwalter (1996) Hand Clin 12(1):13-24; Buckwalter (1995) Iowa Orthop J 15:29-42). As an example, tissue engineered tendons produced in vitro have been shown to be weaker than native adult tendons and would not be expected to withstand rehabilitation after implantation (Garvin et al. (2003) Tissue Eng 9(5):967-979; Calve et al. (2004) Tissue Eng 10(5/6):755-761).
Tendon allografts (i.e., tissue taken from an animal of the same species but not of the same genotype) are useful in ligamentous reconstruction due to their availability and lack of donor site pathology (Poehling et al. (2005) Arthroscopy 21(7):774-85; Cole et al. (2005) Arthroscopy 21(7):786-90). However, these grafts have a micro-architecture that is quite dense and in which appreciable amounts of cellular debris remain, even in freeze-dried grafts. Experience in the preparation and use of allogeneic and xenogeneic (i.e., tissue taken from an animal of a different species) grafts for tissue regeneration has shown that dense grafts with residual cells are difficult to seed in vitro, instigate prolonged inflammatory responses in vivo, and require longer times to incorporate into native tissue and remodel. In clinical practice, this phenomenon has been observed using human allograft tissue and has been shown to result in a delay of remodeling and integration of implanted allografts in comparison to autograft tissue (Jackson et al. (1996) Clin Orthop Rel Res 324:126-33).
Most current strategies for the development of tissue-engineered tendons and ligaments have relied on a construct consisting of cells seeded onto a scaffold. However, the products of recent in vivo and in vitro studies have not produced a construct with sufficient biomechanical strength to withstand immediate rehabilitation after implantation. It is widely accepted that controlled early restoration of activity can promote healing of soft-tissue injuries and that the prolonged rest or immobilization of soft-tissue injuries can result in delayed recovery and permanent loss of function (Buckwalter (1996) Hand Clin 12(1): 13-24; Buckwalter (1995) Iowa Orthop J 15:29-42).
Methods for removing cells from tissues to create a porous bioscaffold are documented (see, e.g., Badylak (2002) Semin Cell Dev Biol 13(5):377-83; U.S. Pat. No. 6,893,666; U.S. Pat. No. 6,962,814; U.S. Pat. No. 6,893,653; U.S. Pat. No. 6,866,686; U.S. Pat. No. 6,753,181; U.S. Pat. No. 6,933,103). A typical method uses a hypo-osmotic solution such as deionized water to burst the cells, followed by a detergent solution, perhaps in combination with an enzyme for disrupting cell adhesion, to remove the cellular debris. A third step is often extraction with a mild basic solution to remove anionic materials such as DNA. The detergent and basic solution are often combined into a single step for convenience. These methods are effective for removal of cellular material without disrupting the native architecture of the extracellular matrix (ECM) or removing many biologically active molecules that mediate cell attachment and growth.
However, because the cells are essentially broken into small fragments and washed away, the porosity of these bioscaffolds is not ideal for repopulating with new cells, whether they are provided via in vitro seeding or by host cell infiltration. This is particularly true if the bioscaffold originated from a dense organ or tissue such as muscle, tendons, ligaments, nerve, blood vessel, cartilage, etc. In fact, most tissues contain a degree of compartmentalization that creates barriers to cell migration. Consequently, seeding cells uniformly throughout such bioscaffolds is extremely difficult, and in some cases impossible.
In addition, ingrowth of seeded or host cells proceeds via a cell-mediated remodeling process that requires considerable time. When a cell seeding approach is used, the subject cells typically attach only to the outside of the bioscaffold and grow outward, away from the bioscaffold.
There remains a need for an improved bioscaffold that addresses these problems and provides an ideal substrate on which cells can grow.